title: Epidermal growth factor (EGF)-nonresponsive variants of normal rat kidney cell line: response to EGF and transforming growth factor-beta.
abstract: Anchorage-independent growth in soft agar of normal rat kidney (NRK) fibroblasts depends on both transforming growth factor-beta (TGF-beta) and epidermal growth factor (EGF) (or TGF-alpha). We have isolated two EGF-nonresponsive cell lines, N-3 and N-9, from chemically mutagenized NRK cells, after selection of mitogen-specific nonproliferative variants in the presence of EGF and colchicine. Saturation binding kinetics with 125I-EGF showed one-half or fewer EGF receptors in N-3 and N-9 than in their parental NRK. Cellular uptake of 2-deoxy-D-glucose was enhanced in all NRK, N-3, and N-9 cell lines by TGF-beta treatment, whereas treatment with EGF significantly enhanced the cellular uptake of the glucose analog in NRK cells, but not in N-3 and N-9 cells. DNA synthesis of NRK during the quiescent state, but not that of N-3 and N-9, was stimulated by EGF. Anchorage-independent growth of N-9 could not be observed even in the presence of both EGF and TGF-beta, whereas that of N-3 was significantly enhanced by TGF-beta alone. EGF stimulated phosphorylation of a membrane protein with molecular size 170 kDa of NRK, but not of N-3, when immunoprecipitates reacting with anti-phosphotyrosine antibody were analyzed. Exposure of NRK cells to EGF increased cellular levels of TGF-beta mRNA, but there appeared little expression of TGF-beta mRNA in N-3 and N-9 cells. Exposure of N-3 cells to EGF or TGF-beta enhanced the secretion of EGF into culture medium, but exposure of NRK or N-9 cells did not. Altered response to EGF of N-3 or N-9 might be related to their aberrant growth behaviors.
authors.affiliation: Department of Biochemistry, Oita Medical School, Japan.
authors.forename: R
authors.initials: R
authors.lastname: Hamanaka
authors.forename: M
authors.initials: M
authors.lastname: Ono
authors.forename: Y
authors.initials: Y
authors.lastname: Kuratomi
authors.forename: H
authors.initials: H
authors.lastname: Mizoguchi
authors.forename: R
authors.initials: R
authors.lastname: Hirai
authors.forename: K
authors.initials: K
authors.lastname: Kohno
authors.forename: M
authors.initials: M
authors.lastname: Kuwano
mesh_terms: D000818:Animals; D002460:Cell Line; D002469:Cell Separation; D004247:DNA / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology*; D066246:ErbB Receptors / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D015870:Gene Expression; D014644:Genetic Variation; D005947:Glucose / Q000493:pharmacokinetics; D007668:Kidney / Q000166:cytology* / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D010766:Phosphorylation; D051381:Rats; D015291:Transforming Growth Factors / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism
chemical_list: D014443:Tyrosine; D004815:Epidermal Growth Factor; D015291:Transforming Growth Factors; D004247:DNA; D066246:ErbB Receptors; D005947:Glucose